T cells are fundamental components of the adaptive immune system, and following exposure to a pathogen, diverse populations of memory T cells persist to provide enhanced protection against re-exposure. While the importance of circulating memory T cells has been appreciated for many years, it is only over the last decade that it has become clear that subsets of memory T cells reside throughout peripheral tissues (known as tissue-resident memory, or T RM) and provide potent, front-line protective immunity. The identification of T RM has resulted in a paradigm shift in the way we need to monitor, target and promote T cell immunity in vaccines, diseases and immunotherapies. However, our understanding of the ontogeny, maintenance, and organization of these cells - their ecology - is lacking, even at a very basic level. For instance, humans retain T-cell immunity to pathogens for years or decades, but it is unclear whether this memory persists as long-lived cells or more dynamically, sustained by self-renewal and/or supplemented by newly generated cells. There are many other open questions; for example, what factors govern the development and maintenance of T RM and how might we manipulate them to boost their numbers? What underlies heterogeneity in their capacity to persist? Do T RM compete with each other, either intra- or inter-clonally? What role does their spatial arrangement play in any competitive dynamics? What are the rules of replacement within T RM niches? In this proposal we will take a multidisciplinary approach to addressing these questions by integrating mathematical and experimental tools in both mouse and human settings. Specifically: (i) We will use a mouse model of influenza infection and quantitative imaging to build a set of validated models of the developmental and homeostatic dynamics of T RM By combining information regarding the time-varying spatial distribution of T RM in the lung, and tracking the responses of pre-existing and newly generated T RM during and after repeat infections, we will refine these models to include competition and spatial niches, and to understand how interaction between T RM influences the rules of replacement within tissues and the durability of T cell memory. (ii) We will use a powerful cell fate-mapping system to model the ontogeny and homeostasis of T RM that are naturally and constitutively produced in multiple tissues across the mouse lifetime, and compare their dynamics to those produced in overt infections. (iii) We will combine a unique human tissue resource at Columbia with a novel application of 14C dating of DNA, a dedicated modeling framework, and quantitative image analysis. This approach will define the contributions of antigen-driven influx and self-renewal to T RM homeostasis, and identify heterogeneity in T RM dynamics, across tissues and ages. In summary, this project will deliver a suite of quantitative tools for defining the life-histories of tissue-specific memory T cells, in both mice and humans, across space and time.